3730xl DNA Analyzer Applied Biosystems technology
User Guide
The instructions in this guide must be followed carefully to achieve the best possible results and avoid delays in processing your request.
Samples that do not comply with the recommendations may be rejected without compensation.
Note that delays in the processing of samples will vary depending on the size of the project. It is recommended to contact the Client Management Office for more information.
- Starting Material
- Requirements
- Primers
- Required containers and sample identification
- Service Request Form and Sample Submission
- Sample Shipment Preparation
- Transmission of results
Starting Material
| Sample Type | Preparation |
|---|---|
| Biological sample requiring extraction | Contact the Client Management Office for information on the Extraction Service |
| Genomic DNA (gDNA) | Commercial kits recommended. In-house protocol: An additional purification step with a commercial kit is required to remove all traces of phenol/chloroform. Elution: 10 mM Tris buffer, pH 8.5, or ultrapure water without nuclease. |
| Bacterial culture | Liquid bacterial culture (~8 hours) |
Requirements
| Sample Type | Volume | Concentration | Recommended method for verifying concentration | Recommended method for verifying purity | Quality criteria to be met |
|---|---|---|---|---|---|
| gDNA | 10 µL of gDNA per fragment to be analyzed | 20 ng/µL | Qubit or equivalent fluorometric method. Note: Spectrophotometric methods such as Nanodrop are much less accurate for measuring DNA concentration. | Nanodrop or other spectrophotometric method. Target optical density ratios: – 260/280 between 1.8 and 2.0 – 260/230 between 2.0 and 2.2. | No degraded or fragmented. |
| Bacterial culture | 1 mL | Target optical density ratios: – OD600 minimum: ~0.4 to 0.6 (growing bacteria) – OD600 maximum: ~1.0 to 1.2 (to avoid overpopulation, which can induce cellular stress and reduce the quality of biomolecule) | Spectrophotometric |
Primers
PCR primer to design
When PCR primers are to be designed, provide the required information such as the rs (Reference SNP) number, the reference sequence, the position in the genome according to UCSC Genome Browser, the species, etc.
PCR primers whose sequence is provided
When the PCR primer sequence is provided by the customer Required containers and sample identification, indicate:
• If these primers are to be ordered and if so, provide the name and sequence of each forward and reverse primer in the Sample Submission Form in the corresponding tab.
• If they are part of the standard primers offered free of charge by the PCR amplification – Sanger Sequencing service, see the following list:
| 27F | 5’ – AGAGTTTGATCMTGGCTCAG – 3’ |
| 1492R | 5’ – TACGGYTACCTTGTTACGACTT – 3’ |
| ITS1F | 5′-CTTGGTCATTTAGAGGAAGTAA-3′ |
| ITS4 | 5′-TCCTCCGCTTATTGATATGC-3′ |
PCR and/or sequencing primers provided
Lorsque les amorces de PCR et/ou de séquençage sont fournies par le client, les règles suivantes sont à respecter:
- The primers must not contain any known SNPs. For any clarification, contact the Client Management Office.
- The primer length must be be between 18 and 24 bases.
- The G/C ratio should be 40-60%.
- PCR product size should be between 250 and 750 bases.
- The primer annealing temperature (Tm) must be above 50°C.
- Primers must be at least 100 bases from the target region to be sequenced.
- Primer design upstream of homo- or heteropolymeric regions (repeats of A, C, G, or T) should be avoided, as these regions are extremely difficult to sequence.
- The Tm and %GC of the primers must be provided.
| Sample Type | Volume | Concentration |
|---|---|---|
| Primer | 5 µL per sample to be analyzed | 20 µM |
Required containers and sample identification
- 48 samples or less: “strip” tubes or 96-well PCR plate.
- 48 samples or more: 96-well PCR plate
The name on the tube or plate must be unique, readable, concise, and identical to the name indicated on the submission form.
Samples must be placed on the plate as indicated on the submission form – Row arrangement mandatory.
This is to avoid any delays in processing the request.
| Sample Type | Container required | Recommended containers | Containers refused |
| Bacterial culture | – 1.5mL tubes All types of 1.5 mL tubes with attached snap caps are accepted. Example: DNA LoBind Tubes, 1.5 mL, PCR clean, colorless; Eppendorf, Cat # 022431021 | ||
| gDNA | – PCR strip tubes of 8 or 12 All types of 200µL PCR tubes in strip of 8 or 12 with a cap are accepted. Example: 0,2 mL PCR strip tubes with 12 well, VWR, Cat # 53509-300 With strip domed caps for 12 well strips, VWR, Cat # 53509-302 – Half-skirt PCR plate All types of clear half-skirt clear 96-well PCR plates are accepted. Example: Thermo-Fast 96 PCR detection plate with flat deck; Life Technologie, Cat# AB1400L Exemple: Thermo-Fast 96 PCR detection plate with flat deck; Life Technologie, Cat# AB1400L – Full-skirt PCR plate All types of clear full-skirt 96-well PCR plates are accepted. – Example: Microseal PCR plates 96-well clear; Bio-Rad, Cat# MSP9601 Clear adhesive film recommended: – Clear adhesive film (Thermo Fisher Scientific, Cat# AB-0558) – 8 or 12-strip caps ensuring a tight seal – Heat sealing to prevent leaks or cross-contamination | – 96-well cell culture plates – No-skirt 96-well PCR plates – Opaques 96-well PCR plates |
Service Request Form and Sample Submission
Click on service request and sample submission to view instructions.
Sample Shipment Preparation
Click on samples shipment preparation to view instructions.
Transmission of results
Sequencing and analysis results, if applicable, are directly accessible online in Nanuq.
An email is sent at the end of the project to inform of the availability of the results.
✔️ Chromatograms and text sequences (FASTA or GenBank format) can be viewed and downloaded locally.
✔️ The SNP detection analysis report contains the following information:
- The position of the SNPs found in the genome
- The identification number (rs) if the SNP is already listed
- The genotype of each sample analyzed
- Whether the SNP is intronic or exonic, and if so, whether it is a synonymous or non-synonymous SNP
- The amino acid change, if applicable.
- The Excel report contains a tab with a tutorial to assist in data interpretation.
✔️ The Excel genotyping analysis report contains the genotype of each sample analyzed.
✔️ The BLAST analysis Word report contains the species and genus, E-value, percent identity, and accession number.
For any questions, please contact the Client Management Office at any time.
Version 1.0_2025-11-06